Structure and Cellular Dynamics of Deinococcus radiodurans Single-stranded DNA (ssDNA)-binding Protein (SSB)-DNA Complexes

• Published in: The Journal of biological chemistry Vol. 287; no. 26; pp. 22123 - 22132
• Main Authors: George, Nicholas P., Ngo, Khanh V., Chitteni-Pattu, Sindhu, Norais, Cédric A., Battista, John R., Cox, Michael M., Keck, James L.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 22.06.2012; American Society for Biochemistry and Molecular Biology
• Subjects: Biochemistry, Molecular Biology; Crystallography, X-Ray; Crystallography, X-Ray - methods; Deinococcus; Deinococcus - metabolism; DNA and Chromosomes; DNA Damage; DNA Damage Response; DNA Recombination; DNA Repair; DNA, Single-Stranded; DNA, Single-Stranded - genetics; DNA-binding Protein; DNA-Binding Proteins; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - metabolism; Escherichia coli; Escherichia coli - metabolism; Ionizing Radiation Resistance; Kinetics; Life Sciences; Microscopy, Electron; Microscopy, Electron - methods; Microscopy, Fluorescence; Microscopy, Fluorescence - methods; Models, Molecular; Oligonucleotides; Oligonucleotides - chemistry; Protein Binding; Protein Structure, Tertiary; Radiation, Ionizing; Recombination, Genetic; Salts; Salts - chemistry; Single-stranded DNA-binding Protein; X-ray Crystallography; DNA Damage Response; X-ray Crystallography; DNA Repair; DNA-binding Protein; DNA Recombination; Ionizing Radiation Resistance; Single-stranded DNA-binding Protein
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.367573

The single-stranded DNA (ssDNA)-binding protein from the radiation-resistant bacterium Deinococcus radiodurans (DrSSB) functions as a homodimer in which each monomer contains two oligonucleotide-binding (OB) domains. This arrangement is exceedingly rare among bacterial SSBs, which typically form homotetramers of single-OB domain subunits. To better understand how this unusual structure influences the DNA binding and biological functions of DrSSB in D. radiodurans radiation resistance, we have examined the structure of DrSSB in complex with ssDNA and the DNA damage-dependent cellular dynamics of DrSSB. The x-ray crystal structure of the DrSSB-ssDNA complex shows that ssDNA binds to surfaces of DrSSB that are analogous to those mapped in homotetrameric SSBs, although there are distinct contacts in DrSSB that mediate species-specific ssDNA binding. Observations by electron microscopy reveal two salt-dependent ssDNA-binding modes for DrSSB that strongly resemble those of the homotetrameric Escherichia coli SSB, further supporting a shared overall DNA binding mechanism between the two classes of bacterial SSBs. In vivo, DrSSB levels are heavily induced following exposure to ionizing radiation. This accumulation is accompanied by dramatic time-dependent DrSSB cellular dynamics in which a single nucleoid-centric focus of DrSSB is observed within 1 h of irradiation but is dispersed by 3 h after irradiation. These kinetics parallel those of D. radiodurans postirradiation genome reconstitution, suggesting that DrSSB dynamics could play important organizational roles in DNA repair. The dimeric D. radiodurans single-stranded DNA-binding protein (DrSSB) has poorly defined DNA-binding and cellular mechanisms. DrSSB binds ssDNA analogously to tetrameric bacterial SSBs and is regulated in response to ionizing radiation. The DrSSB ssDNA mechanism is conserved with other SSBs, and the protein is dynamically localized during DNA repair. The findings suggest a central role for DrSSB in DNA repair.